The unique role of dietary l-arginine in the acceleration of peritoneal macrophage sensitivity to bacterial endotoxin

Abstract

It is known that cells and organisms can indirectly "sense" changes in L-arginine availability via changes in the activity of various metabolic pathways. However, the mechanism(s) by which genes can be directly regulated by L-arginine in mammalian cells have not yet been elucidated. We investigated the effect of L-arginine in the in vivo model of peritoneal inflammation in mice and in vitro in RAW 264.7 macrophages. A detailed analysis of basic physiological functions and selected intracellular signaling cascades revealed that L-arginine is crucial for the acceleration of macrophage activation by bacterial lipopolysaccharide. L-arginine increased the production of reactive oxygen species, nitric oxide, release of Ca(2+), as well as inducible nitric oxide synthase expression. Interestingly, the effect of L-arginine on macrophage activation was dependent on the phosphorylation of mitogen-activated protein kinases and activity of phospholipase C. In RAW 264.7 cells, L-arginine was shown to modulate the response of macrophages toward lipopolysaccharide via the activation of G-protein-coupled receptors. According to our data, we concluded that L-arginine availability plays a key role in the initiation of intracellular signaling pathways that trigger the lipopolysaccharide-induced inflammatory responses in murine macrophages. Although macrophages are partially stimulated in the absence of extracellular L-arginine, the presence of this amino acid significantly accelerates the sensitivity of macrophages to bacterial endotoxin.

The crucial role of l-arginine in macrophage activation: What you need to know about it

“…Therefore, the most interesting and promising candidate for L-arginine effect on macrophages is the GPRC6A receptor, the activation of which was found to be responsible for the L-arginine-mediated NO signaling in endothelial cells (Christiansen et al, 2007;Joshi et al, 2007). A similar pathway was documented for the L-arginine-mediated MAPK signaling in macrophages (Pekarova et al, 2013b). The activation of GPRC6A by extracellular L-arginine led to a significant increase in the LPS-stimulated macrophage NO synthesis.…”

“…Importantly, this also involves the activation of the innate , 1996). Importantly, it was demonstrated that the decrease in L-arginine concentration, occurring during inflammatory processes, could be associated with a significant reduction of NO levels released from activated macrophages (El-Gayar et al, 2003;Pekarova et al, 2011;Pekarova et al, 2013b). This phenomenon, which is crucially involved in the deregulation of innate immune responses, was described as the "L-arginine paradox."…”

“…This phenomenon, which is crucially involved in the deregulation of innate immune responses, was described as the "L-arginine paradox." It refers to the dependence of cellular NO production on extracellular L-arginine concentration, despite the theoretical saturation of NOS enzymes with intracellular L-arginine, which can be endogenously synthesized in macrophages (Boger et al, 2004;Lee et al, 2003;Nicholson et al, 2001;Pekarova et al, 2011;Pekarova et al, 2013b;Wu and Brosnan, 1992). It was demonstrated that L-arginine deficiency transiently reduces not only the production of NO by iNOS due to the lack of substrate for the enzyme but also the synthesis and, to a lesser degree, the stability of iNOS in macrophages and other cell types (El-Gayar et al, 2003;Jousee et al, 2004;Kagemann et al, 2007, Lee et al, 2003.…”

Bone marrow-derived macrophages exclusively expressed caveolin-2: The role of inflammatory activators and hypoxia

“…Cells were transiently transfected with control siRNA and siRNA against caveolin-2 (all siRNA constructs were purchased from Santa Cruz Biotechnology, USA) using an electroporation system (Gene Pulser II, Bio-Rad Laboratories) as described previously (Pekarova et al, 2013a). There was no change in cell viability when compared macrophages transfected with control siRNA and macrophages transfected with siRNA against caveolin-2 (data not shown).…”

“…All experiments were conducted in C57BL/6J mice (males, 25-30 g, 12-14 weeks old) (Masaryk University, Brno, Czech Republic) housed in a temperature-controlled animal facility with 12 h light-dark cycle and free access to rodent chow. BMDMs and peritoneal macrophages (PMs) (used as a positive control) were isolated from mice according to standard protocols, described previously in more details (Pekarova et al, 2013a;Rudolph et al, 2010). BMDMs were grown in Dulbecco's Modified Eagle Media (DMEM), which were supplemented with culture medium from CCL-1 cells containing macrophage-colony stimulating factor (M-CSF) (ATCC, Manassas, VA, USA), 20% of fetal bovine serum (FBS, low endotoxin; PAA, Pasching, Austria), and 1% gentamycin.…”